بررسی اثر مهاری سیلیمارین بر فرایند فیبریلاسیون پروتئین لیزوزیم سفیده تخم مرغ

نوع مقاله: مقاله پژوهشی

نویسندگان

1 دانشجوی کارشناسی ارشد بیوشیمی، دانشکده علوم زیستی، دانشگاه تحصیلات تکمیلی علوم پایه زنجان، زنجان، ایران

2 دانشکده علوم زیستی، دانشگاه تحصیلات تکمیلی علوم پایه زنجان، جاده گاوازنگ، زنجان، ایران

چکیده

بسیاری از مطالعات انجام شده در سالهای اخیر نشان میدهد که ترکیبات حلقوی پلی فنلی که به وفور در بسیاری از گیاهان وجود دارند قادر به مهار تشکیل فیبریلهای آمیلوئیدی هستند. با اینحال مکانیسم دقیقی که بدان وسیله این ترکیبات طبیعی روند تشکیل تجمعات آمیلوئیدی را تحت تاثیر قرار میدهند بطور کامل شناسائی نشده است. گیاه خارمریم (Silybum Marianum) از جمله گیاهان دارویی میباشد که دارای خواص درمانی بسیار زیادی است. عصاره حاصل از دانۀ این گیاه که تحت عنوان سیلیمارین نامیده میشود غنی از ترکیبات فلاونولیگنانی و فلاونوئیدی است. در این مطالعه، با استفاده از طیف وسیعی از تکنیکها شامل نشر فلورسانس نشانگرهای تیوفلاوین T و Nile red، سنجش میزان جذب قرمز کنگو، طیف سنجی دورنگ‌ نمایی دورانی، و میکروسکوپ نیروی اتمی، توانایی سیلیمارین در مهار تشکیل تجمعات آمیلوئیدی پروتئین لیزوزیم سفیده تخم مرغ مورد بررسی قرار گرفت. نتایج ما نشان داد که سیلیمارین، بطور وابسته به غلظت، باعث مهار تشکیل تجمعات آمیلوئیدی میشود. نتایج حاصل از میکروسکوپ نیروی اتمی نیز دلالت بر تشکیل فیبریلهای آمیلوئیدی در نمونه های کنترل داشت، حال آنکه نمونه های پروتئینی انکوبه شده در حضور سیلیمارین فاقد فیبریلهای آمیلوئیدی بوده و مقادیر جزئی از تجمعات پروتئینی بدون شکل در آنها مشاهده شد. براساس نتایج حاصل پیشنهاد میشود که ممانعت از برهمکنشهای آبگریز بین گونه های پروتئینیِ بطور نسبی بازشده، مکانیسمی است که بدان وسیله سیلیمارین باعث مهار تشکیل تجمعات آمیلوئیدی در پروتئین لیزوزیم میشود. با اینحال مطالعات و آزمایشات بیشتری برای تعیین دقیق مکانیسم عمل سیلیمارین مورد نیاز میباشد.

کلیدواژه‌ها


عنوان مقاله [English]

A Study on the Inhibitory Effects of Silymarin on Amyloid Fibrillation of Hen Egg White Lysozyme

نویسندگان [English]

  • Mohsen Mahdavimehr 1
  • Ali Akbar Meratan 2
1 M.Sc. student in Biochemistry, Department of Biological Sciences, Institute in Advanced Studies in Basic Sciences (IASBS), Zanjan, Iran
2 Department of Biological Sciences, Institute of Advanced Studies in Basic Sciences (IASBS), Zanjan, Iran.
چکیده [English]

An increasing number of studies have demonstrated that polyphenols, compounds frequently occurring in many herbs with antioxidant properties, prevent amyloid fibril formation. However, the mechanisms by which these natural molecules modulate the protein aggregation process are poorly understood. Silybum Marianum is one of the medicinal plants with a wide range of health benefits. Silymarin, extract of the seeds of Silybum Marianum, contains a mixture of flavonolignans and a flavonoid. In the present study, using a range of techniques including Thioflavin T and Nile red fluorescence assays, Congo red binding assay, Circular Dichroism spectroscopy, and Atomic Force Microscopy the efficacy of Silymarin on the inhibition of Hen Egg White Lysozyme (HEWL) fibril formation was investigated. Obtained results demonstrated that Silymarin effectively inhibits fibrillogenesis of HEWL in a concentration-dependent manner. AFM images indicated typical fibrillation in the control solutions, while in samples incubated in the presence of Silymarin extensive inhibition of HEWL fibrillation and amorphous aggregates formation was observed. Based on obtained results, we suggest that preventing of hydrophobic interactions between HEWL amyloidogenic prefibrillar species is the mechanism by which Silymarin inhibits amyloid fibril formation by HEWL. However, additional studies are needed to elucidate the detailed mechanisms involved.

کلیدواژه‌ها [English]

  • amyloid
  • Polyphenol
  • Protein Aggregation
  • Silybum marianum
  • silymarin
[1]. Azami-Movahed, M., Shariatizi, S., Sabbaghian, M., Ghasemi, A., EbrahimHabibi, A., Nemat-Gorgani, M. 2013. Heme binding site in apomyoglobin may be effectively targeted with small molecules to control aggregation. -Int. J. Biochem. Cell Biol. 45(2): 299-307.

[2]. Bartolini, M., Andrisano, V. 2010. Strategies for the inhibition of protein aggregation in human diseases.- ChemBioChem. 11(8): 1018–1035.

[3]. Borana, M.S., Mishra, P., Pissurlenkar, R.R., Hosur, R.V., Ahmad, B. 2014. Curcumin and kaempferol prevent lysozyme fibril formation by modulating aggregation kinetic parameters. -Biochim. Biophys. Acta. 1844(3): 670–680.

[4]. Bouma, B., Kroon-Batenburg, L.M., Wu, Y.P., Brünjes, B., Posthuma, G., Kranenburg, O., de Groot, P.G., Voest, E.E., Gebbink, M.F. 2003. Glycation induces formation of amyloid cross-beta structure in albumin. -J. Biol. Chem. 278 (43): 41810-9.

[5]. Bucciantini, M., Giannoni, E., Chiti, F., Baroni, F., Formigli, L., Zurdo, J., Taddei, N., Ramponi, G., Dobson, C.M., Stefani, M. 2002. Inherent toxicity of aggregates implies a common mechanism for protein misfolding diseases.- Nature, 416(6880): 507–511.

[6]. Cheng, B., Gong, H., Li, X., Sun, Y., Zhang, X., Chen, H., Liu, X., Zheng, L., Huang, K. 2012. Silibinin inhibits the toxic aggregation of human islet amyloid polypeptide. -Biochem Biophys Res Commun. 419(3): 495-9.

[7]. Cohen, F.E., Kelly, J.W. 2003. Therapeutic approaches to protein-misfolding diseases. -Nature. 426(6968): 905–909.

[8]. Cohen, T., Frydman-Marom, A., Rechter, M., Gazit, E. 2006. Inhibition of amyloid fibril formation and cytotoxicity by hydroxyindole derivatives. -Biochemistry 45(15): 4727–4735.

[9]. De Felice, F.G., S.T. Ferreira, S.T. 2002. Beta-Amyloid production, aggregation, and clearance as targets for therapy in Alzheimer’s disease. -Cell. Mol. Neurobiol. 22(5-6): 545–563.

[10]. De Felice, F.G., Houzel, J.C., GarciaAbreu, J., Louzada Jr., P.R., Afonso, R.C. Meirelles, M.N., Lent, R., Neto, V.M., Ferreira, S.T. 2001. Inhibition of Alzheimer’s disease beta-amyloid aggregation, neurotoxicity, and in vivo deposition by nitrophenols: implications for Alzheimer’s therapy. -FASEB J. 15(7): 1297–1299.

[11]. Ebrahimi, A., Schluesener, H. 2012. Natural polyphenols against neurodegenerative disorders: potentials and pitfalls. -Ageing Res. Rev. 11(2): 329–345.

[12]. Ferreira, N., Saraiva, M.J., Almeida, M.R. 2011. Natural polyphenols inhibit different steps of the process of transthyretin (TTR) amyloid fibril formation. -FEBS Lett. 585(15): 2424–2430.

[13]. Gazit, E. 2002. A possible role for pistacking in the self-assembly of amyloid fibrils. -FASEB J. 16(1): 77–83.

[14]. Gazit, E. 2005. Mechanisms of amyloid fibril self-assembly and inhibition. Model short peptides as a key research tool.- FEBS J. 272(23): 5971–5978.

[15]. Gazova, Z., Siposova, K., Kurin, E., Mucaji, P., Nagy, M. 2013. Amyloid aggregation of lysozyme: the synergy study of red wine polyphenols. -Proteins. 81(6): 994–1004.

[16]. Ghobeh, M., Ahmadian, S., Meratan, A.A., Ebrahim-Habibi, A., Ghasemi, A., Shafizadeh, M., Nemat-Gorgani, M. 2014. Interaction of Aβ(25–35) fibrillation products with mitochondria: effect of small-molecule natural products. Biopolymers. 102(6): 473–486.

[17]. Goldberg, M.E., Rudolph, R., Jaenicke, R. 1991. A kinetic study of the competition between renaturation and aggregation during the refolding of denatured-reduced egg white lysozyme. -Biochemistry. 30(11): 2790–2797.

[18]. Hafner-Bratkovic, I., Gaspersic, J., Smid, L.M., Bresjanac, M., Jerala, R. 2008. Curcumin binds to the alpha-helical intermediate and to the amyloid form of prion protein –a new mechanism for the inhibition of PrP (Sc) accumulation. -J. Neurochem. 104(6): 1553–1564. 

[19]. Härd, T., Lendel, C. 2012. Inhibition of amyloid formation. -J. Mol. Biol. 421(4-5): 441–465.

[20]. Kayed, R., Head, E., Thompson, J.L., McIntire, T.M., Milton, S.C., Cotman, C.W., Glabe, C.G. 2003. Common structure of soluble amyloid oligomers implies common mechanisms of pathogenesis. -Science. 300(5618): 486– 489.

[21]. Klunk, W.E., Pettegrew, J.W., Abraham, D.J. 1989. Quantitative evaluation of Congo red binding to amyloid-like proteins with a beta-pleated sheet conformation. -J. Histochem. Cytochem. 37(8): 1273–1281.

[22]. Krebs, M.R., Wilkins, D.K., Chung, E.W., Pitkeathly, M.C., Chamberlain, A.K., Zurdo, J., Robinson, C.V., Dobson, C.M. 2000. Formation and seeding of amyloid fibrils from wild-type hen lysozyme and a peptide fragment from the β-domain. J.Mol. Biol. 300(3): 541–549.

[23]. Masuda, M., Suzuki, N., Taniguchi, S., Oikawa, T., Nonaka, T., Iwatsubo, T., Hisanaga, S., Goedert, M., Hasegawa, M. 2006. Small molecule inhibitors of alphasynuclein filament assembly. Biochemistry. 45(19): 6085–6094.

[24]. Meratan, A.A., Ghasemi, A., NematGorgani, M. 2011. Membrane integrity and amyloid cytotoxicity: a model study involving mitochondria and lysozyme fibrillation products. -J. Mol. Biol. 409(5): 826–838.

[25]. Mishra, R., Sjölander, D., Hammarström, P. 2011. Spectroscopic characterization of diverse amyloid fibrils in vitro by the fluorescent dye Nile red. -Mol Biosyst. 7(4): 1232-40.

[26]. Mishra, R., Sellin, D., Radovan, D., Gohlke, A., Winter, R. 2009. Inhibiting islet amyloid polypeptide fibril formation by the red wine compound resveratrol. ChemBioChem. 10(3): 445–449.

[27]. Murata, N., Murakami, K., Ozawa, Y., Kinoshita, N., Irie, K., Shirasawa, T., Shimizu, T. 2010. Silymarin attenuated the amyloid β plaque burden and improved behavioral abnormalities in an Alzheimer's disease mouse model. -Biosci Biotechnol Biochem. 74(11): 2299-306.

[28]. Ono, K., Yamada, M. 2006. Antioxidant compounds have potent anti-fibrillogenic and fibril-destabilizing effects for alphasynuclein fibrils in vitro. -J. Neurochem. 97(1): 105–115.

[29]. Pepys, M.B. Hawking, P.N. Booth, D.R., Vigushin, D.M., Tennent, G.A., Soutar, A.K., Totty, N., Nguyen, O., Blake, C.C., Terry, C.J., Feest, T.G., Zalin, A.M., Hsuan, J.J. 1993. Human lysozyme gene mutations cause hereditary systemic amyloidosis. -Nature. 362(6420): 553–557.

[30]. Porat, Y., Abramowitz, A., Gazit, E. 2006. Inhibition of amyloid fibril formation by polyphenols: structural similarity and aromatic interactions as a common inhibition mechanism. -Chem. Biol. Drug Des. 67(1): 27–37.

[31]. Polyak, S.J., Ferenci, P., Pawlotsky, J.M. 2013. Hepatoprotective and antiviral functions of silymarin components in hepatitis C virus infection. -Hepatology. 57(3): 1262-71.

[32]. Ramasamy, K., Agarwal, R. 2008. Multitargeted therapy of cancer by silymarin. -Cancer Lett. 269(2): 352-62.

[33]. Shariatizi, S., Meratan, A.A., Ghasemi, A., Nemat-Gorgani, M. 2015. Inhibition of amyloid fibrillation and cytotoxicity of lysozyme fibrillation products by polyphenols. -Int. J. Biol. Macromol. 80: 95–106.

[34]. Soto, C. 1999. Plaque busters: strategies to inhibit amyloid formation in Alzheimer’s disease. -Mol. Med. Today. 5(8): 343–350.

[35]. Soto, C. 2003. Unfolding the role of protein misfolding in neurodegenerative diseases. -Nat. Rev. Neurosci. 4(1): 49–60.

[36]. Uversky, V.N., Fink, A.L. 2004. Conformational constraints for amyloid fibrillation: the importance of being unfolded.- Biochim. Biophys. Acta. 1698(2): 131–153.

[37]. Wang, J.B., Wang, Y.M., Zeng, C.M. 2011. Quercetin inhibits amyloid fibrillation of bovine insulin and destabilizes preformed fibrils. -Biochem. Biophys. Res. Commun. 415(4): 675–679.

[38]. Zeng, H., Miao, M., Yang, R., Qu, L. 2017. Effect of silybin on the fibrillation of hen egg white lysozyme. -J. Mol. Recognit. 30(1): e2566.